The development of DNA assay in a miniaturized device is an essential element for point-of-care testing (POCT) like decentralized healthcare, pathogen identification, and environmental monitoring. With the advanced technology in micro-fabrication as well as the compatibility of electrical/electrochemical (EC) DNA detection in the micro-fabricated platform, an integrated micro-device with multiple functionalities can be produced to perform routine DNA analysis. In this thesis, a novel EC DNA assay in a silicon/glass-based micro-device is developed....[ Read more ]

The development of DNA assay in a miniaturized device is an essential element for point-of-care testing (POCT) like decentralized healthcare, pathogen identification, and environmental monitoring. With the advanced technology in micro-fabrication as well as the compatibility of electrical/electrochemical (EC) DNA detection in the micro-fabricated platform, an integrated micro-device with multiple functionalities can be produced to perform routine DNA analysis. In this thesis, a novel EC DNA assay in a silicon/glass-based micro-device is developed.

The thesis starts with the development of a novel sample-to-answer DNA assay in a biochip, which is equipped with a fabricated thermal system and electrodes. In contrast to most reported designs which compose of numerous reaction chambers interconnected with microfluidic channels, the assay performs thermal cell lysis, magnetic particles-assisted genomic DNA extraction, polymerase chain reaction, and EC DNA detection within a single reaction chamber. Coelectropolymerization of pyrrole and pyrrole-DNA probe for selective electrode modification will be studied for multiplex DNA assay. As individual module has been studied in the research group, the presented work focuses on the issues concerning the compatibilities among different components in the assay in the biochip system. It is believed that the novel single reaction chamber design for sample-to-answer DNA analysis will provide a simplified solution for decentralized DNA detection.

The real-time PCR features a simple assay with high accuracy and sensitivity. However, the complicated optics makes it too bulky and expensive for POCT. The second part of thesis focuses on the development of EC real-time PCR (ERT-PCR). The proposed scheme involves ferrocene-based reporter and solid phase primer extension for simultaneous PCR and detection. Issues concerning redox signal accumulation, probe immobilization chemistry, electrode surface passivation, electrical scanning scheme and controlling factors (i.e. enzyme and DNA probe availability) will be studied. Moreover, the developed ERT-PCR will be implemented in a biochip system and applied in qualitative and quantitative HBV DNA detection. It is believed that the novel ERT-PCR biochip presented in this thesis is a great step towards the realization of a portable EC-based DNA sensor for POCT applications.